CN104756872A - Seed sterilization method for horseradish tree tissue culture - Google Patents
Seed sterilization method for horseradish tree tissue culture Download PDFInfo
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- CN104756872A CN104756872A CN201510209011.5A CN201510209011A CN104756872A CN 104756872 A CN104756872 A CN 104756872A CN 201510209011 A CN201510209011 A CN 201510209011A CN 104756872 A CN104756872 A CN 104756872A
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Abstract
The invention belongs to the field of tissue culture and particularly relates to a seed (pre-treatment) sterilization method for horseradish tree tissue culture. The seed sterilization method for horseradish tree tissue culture comprises the steps that paper wings and brown shells of seeds are removed, and then sterilization is conducted on the seeds by means of 75% alcohol and 0.1% mercury bichloride. The seed sterilization method for horseradish tree tissue culture is provided for solving the problems that an existing sterilization method is complicated and time consumption is high. Compared with an existing seed sterilization method for horseradish tree tissue culture, the seed sterilization method for horseradish tree tissue culture has the advantages that the sterilization time is short, the contamination rate is small, the germination rate is high, and a high emergence rate is guaranteed.
Description
Technical field
The invention belongs to field of tissue culture, be specially seed (pretreatment) sterilization method in a kind of Moringa tissue cultures.
Background technology
Moringa is Moringaceae Moringa plant, is rich in the various nutrient elements such as protein, calcium, iron, potassium, Va, Vc.Moringa is not only edible, and is also widely used in health care.Along with continually developing of Moringa function, the planting scale of Moringa is also in continuous expansion.
Along with plant tissue culture technique is more and more ripe, be now applied in various plants and carried out Fast-propagation.In Plant Tissue Breeding, obtain the first step that aseptic seedling is success of the test, and aseptic seedling is generally obtained by seed, therefore, effective sterilization method of seed seems particularly important.
At present, the sterilization method about Moringa seed in Moringa tissue cultures has been reported (Luo Yunxia etc., 2007), but its sterilization method is more loaded down with trivial details, consuming time longer, adds the toxic to seed.
Summary of the invention
For the deficiency of above-mentioned technology, the invention provides a kind of seed disinfection method in Moringa tissue cultures.
The present invention is achieved through the following technical solutions:
A seed disinfection method in Moringa tissue cultures, is characterized in that: before seed disinfection, the papery assistant of seed and brown shell is removed, then utilizes the alcohol of 75% and the mercuric chloride disinfection of 0.1%.
This method comprises the steps:
(1) select the Moringa seed of full seed, papery assistant and brown shell are removed;
(2) with aseptic water washing;
(3) with 75% alcohol disinfecting;
(4) sterilize with the mercuric chloride of 0.1%;
(5) finally sterilisation step is completed with aseptic water washing.
Aseptic water washing 1-2 time is used in step (2).
In step (3), the alcohol disinfecting time is 30s.
Add 1-2 in step (4) and drip Tween-20.
Step (4) the middle time with the mercuric chloride sterilization of 0.1% is 7-15min, is excellent with 10min.
Aseptic water washing 5-6 time is used in step (5).
In Moringa tissue cultures, existing seed disinfection method, seed need through the step such as running water, immersion, and disinfectant to disinfect the time longer; In the present invention, seed is without the need to through the step such as running water, immersion, thus shortens the seed treatment time, and when disinfectant is disinfected, disinfecting time shortens simultaneously, reduces the toxic action of disinfectant to seed.The method of the invention, compared with the seed disinfection method in existing Moringa tissue cultures, has the advantage that the seed treatment time is short, disinfecting time is short, pollution rate is little, germination rate is high, ensure that higher emergence rate.
Embodiment
Below in conjunction with embodiment, the present invention is further described, but the present invention is not limited thereto.The present embodiment all with the Moringa seed purchased from India, Xiamen Industrial Co., Ltd. for test material.
Application Example one
The present embodiment adopts three kinds of processing modes to carry out pretreatment to seed, and the mercuric chloride disinfecting time of having carried out 0.1% is to the influence research of Moringa seed Disinfection Effect.
One, Moringa seed preprocess method design:
seed removes papery assistant;
seed removes papery assistant and brown shell;
undressed seed.
Seed disinfection method: each process 100 seeds, aseptic water washing 2 times used by superclean bench, then uses the alcohol disinfecting 30s of 75%, then add 2 Tween-20s with the mercuric chloride sterilization 10min(of 0.1%), finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Two, the mercuric chloride disinfecting time of 0.1% is on the impact of Moringa seed Disinfection Effect
Moringa seed disinfecting time designs:
the mercuric chloride sterilization 7min of 0.1%;
the mercuric chloride sterilization 10min of 0.1%;
the mercuric chloride sterilization 15min of 0.1%.
Seed disinfection method: each process 100 seeds, the seed removing papery assistant and brown shell is used aseptic water washing 2 times on superclean bench, then the alcohol disinfecting 30s of 75% is used, disinfect (adding 2 Tween-20s) with the mercuric chloride of 0.1% again, finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Three, Moringa Aseptic seedling culture
By the Moringa seed after disinfecting, be inoculated on MS medium, be placed in culturing room and cultivate, cultivation temperature (25 ± 2) DEG C, intensity of illumination 1500-2000lx, light application time 12h/d.
Cultivate the Disinfection Effect observing seed for 3 days afterwards, when cultivating 15 days, observe pollution rate, the germination rate of statistics seed.
Results and analysis is as follows:
Table 1 pretreating method for seed germinates and the impact of Disinfection Effect on Moringa seed
As shown in Table 1, the germination rate of process 2 is 86%, pollution rate is 0%, is obviously better than process 1,3, it can thus be appreciated that the Disinfection Effect of preprocess method to seed of process 2 has positive role, is realize high germination rate, the comparatively ideal method of low stain rate.
Table 2 disinfecting time is germinateed and the impact of Disinfection Effect on Moringa seed
As shown in Table 2, process 1,2,3 can reach good Disinfection Effect, and the pollution rate of process 2,3 is 0%, but the germination rate of process 3 is lower than process 1,2.It can thus be appreciated that the length of disinfecting time affects the germination rate of seed, to process the mercuric chloride sterilization 10min of 2:(0.1%) for good.
Application Example two
Embodiment 1
With the Moringa seed purchased from India, Xiamen Industrial Co., Ltd. for test material, the seed removing papery assistant is used aseptic water washing 2 times on superclean bench, then the alcohol disinfecting 30s of 75% is used, disinfect 10min(add 2 Tween-20s with the mercuric chloride of 0.1% again), finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Embodiment 2
With the Moringa seed purchased from India, Xiamen Industrial Co., Ltd. for test material, the seed removing papery assistant and brown shell is used aseptic water washing 2 times on superclean bench, then the alcohol disinfecting 30s of 75% is used, disinfect 10min(add 2 Tween-20s with the mercuric chloride of 0.1% again), finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Embodiment 3
With the Moringa seed purchased from India, Xiamen Industrial Co., Ltd. for test material, undressed seed is used aseptic water washing 2 times on superclean bench, then the alcohol disinfecting 30s of 75% is used, disinfect 10min(add 2 Tween-20s with the mercuric chloride of 0.1% again), finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Embodiment 4
With the Moringa seed purchased from India, Xiamen Industrial Co., Ltd. for test material, the seed removing papery assistant and brown shell is used aseptic water washing 2 times on superclean bench, then the alcohol disinfecting 30s of 75% is used, disinfect 7min(add 2 Tween-20s with the mercuric chloride of 0.1% again), finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Embodiment 5
With the Moringa seed purchased from India, Xiamen Industrial Co., Ltd. for test material, the seed removing papery assistant and brown shell is used aseptic water washing 2 times on superclean bench, then the alcohol disinfecting 30s of 75% is used, disinfect 10min(add 2 Tween-20s with the mercuric chloride of 0.1% again), finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Embodiment 6
With the Moringa seed purchased from India, Xiamen Industrial Co., Ltd. for test material, the seed removing papery assistant and brown shell is used aseptic water washing 2 times on superclean bench, then the alcohol disinfecting 30s of 75% is used, disinfect 15min(add 2 Tween-20s with the mercuric chloride of 0.1% again), finally use aseptic water washing 5-6 time, be inoculated in MS medium.
Claims (7)
1. the seed disinfection method in Moringa tissue cultures, is characterized in that: the method comprises the following steps:
(1) select the Moringa seed of full seed, papery assistant and brown shell are removed;
(2) with aseptic water washing;
(3) with 75% alcohol disinfecting;
(4) sterilize with the mercuric chloride of 0.1%;
(5) finally sterilisation step is completed with aseptic water washing.
2. the seed disinfection method in Moringa tissue cultures according to claim 1, is characterized in that: use aseptic water washing 1-2 time in step (2).
3. the seed disinfection method in Moringa tissue cultures according to claim 1, is characterized in that: in step (3), the alcohol disinfecting time is 30s.
4. the seed disinfection method in Moringa tissue cultures according to claim 1, is characterized in that: add 1-2 in step (4) and drip Tween-20.
5. the seed disinfection method in Moringa tissue cultures according to claim 1, is characterized in that: step (4) the middle time with the mercuric chloride sterilization of 0.1% is 7-15min.
6. the seed disinfection method in Moringa tissue cultures according to claim 5, is characterized in that: step (4) the middle time with the mercuric chloride sterilization of 0.1% is 10min.
7. the seed disinfection method in Moringa tissue cultures according to claim 1, is characterized in that: use aseptic water washing 5-6 time in step (5).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201510209011.5A CN104756872A (en) | 2015-04-29 | 2015-04-29 | Seed sterilization method for horseradish tree tissue culture |
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| CN201510209011.5A CN104756872A (en) | 2015-04-29 | 2015-04-29 | Seed sterilization method for horseradish tree tissue culture |
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| CN201510209011.5A Pending CN104756872A (en) | 2015-04-29 | 2015-04-29 | Seed sterilization method for horseradish tree tissue culture |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105961197A (en) * | 2016-05-09 | 2016-09-28 | 华南农业大学 | High efficiency regeneration method of Moringa oleifera |
| CN106718941A (en) * | 2017-02-15 | 2017-05-31 | 钦州市林业科学研究所 | A kind of construction method of Moringa regenerating system |
-
2015
- 2015-04-29 CN CN201510209011.5A patent/CN104756872A/en active Pending
Non-Patent Citations (1)
| Title |
|---|
| 张婧: "辣木组织培养及有效成分分析", 《中国优秀硕士学位论文全文数据库》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105961197A (en) * | 2016-05-09 | 2016-09-28 | 华南农业大学 | High efficiency regeneration method of Moringa oleifera |
| CN106718941A (en) * | 2017-02-15 | 2017-05-31 | 钦州市林业科学研究所 | A kind of construction method of Moringa regenerating system |
| CN106718941B (en) * | 2017-02-15 | 2019-04-19 | 钦州市林业科学研究所 | A kind of construction method of Moringa regenerating system |
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Application publication date: 20150708 |